Scalable Preparation of Synthetic Mucins via Nucleophilic Ring-Opening Polymerization of Glycosylated N-Carboxyanhydrides

A method for the preparation of glycosylated polypeptides via the nucleophilic ring-opening polymerization of a glycosylated N-carboxyanhydride (NCA) monomer is reported. The synthesis of 2,3,4,6-tetraacetyl-beta-galactose-threonine Ncarboxyanhydride (beta-AcO-Gal-Thr-NCA) monomer in five steps with an 8% overall yield is described, and the single-crystal X-ray structure is provided. The effects of a series of Ni0-based organometallic initiators, nucleophilic amine initiators, cocatalysts, and solvents on the polymerization were explored. The kinetics of the three most promising conditions were studied in greater detail. The conditions that provided the highest yield, low polydispersity (??), and excellent control over the degree of polymerization (X?? n), while being amenable to gram-scale reactions, involved the nucleophilic initiator lithium hexamethyldisilazide (LiHMDS) and the cocatalyst 1,3-bis(2-hydroxyhexafluoroisopropyl) benzene (HFAB) in the nonpolar solvent CH2Cl2. A detailed analysis of this polymerization revealed that two propagation reactions proceed simultaneously, although at substantially different rates. These optimized polymerization conditions provide a route toward the synthesis of polymers that mimic the structures and properties of the highly glycosylated proline, threonine, and serine (PTS) domains of secreted mucin proteins.